Abstract E-cigarette (E-cig) use has rapidly proliferated worldwide. It is speculated that E-cig use may exceed conventional cigarette use within 10 years. Cigarette smoking is the leading preventable cause of death and disability. As E-cigs are a relatively new product, studies are needed to determine their long-term detrimental effects, some of which may be similar to the long-term detrimental effects of nicotine. The lack of an appropriate method for E-cig delivery to animals and lack of an appropriate animal model are major barriers of the research field. We propose to develop a system designed for E-cig exposure to rodent models. We propose that E-cig exposure using our system can generate clinically relevant animal models with nicotine levels/PK comparable to those of human E-cig users for research of E-cig effects on human health, and allow investigators to uncover potential detrimental and beneficial effects related to E-cigs. Aim 1. To design and make a prototype for a product of efficient E-cig aerosol generation and exposure system delivering E- cig aerosol to rodent(s) through inhalation with characteristics equivalent to those inhaled by human E-cig users. This system includes hardware and software that control the number of E-cigs and timing of activation so as to regulate the dose of aerosol exposure. This device has variable E-cig holders for different sizes of E-cigs available in the market, and generates aerosol for testing different brands of E-cigs of investigators' choice. We will characterize the E-cig aerosol by measuring the aerosol particle size distribution and mass concentration in the breathing zone of the rodent exposure chamber to verify that the aerosol characteristics are equivalent to those in mainstream E-cigs that human E-cig smokers inhale. Aim 2. To validate this E-cig aerosol generation and exposure system with acute and chronic animal experiments to show that it produces E-cig animal models with nicotine PK comparable to that in human E-cig users. We will measure plasma nicotine and cotinine levels at a series of time points during and after rats or mice are exposed to E-cig aerosol in a free-moving or nose-only chamber. We will optimize the parameters (e.g., number of E-cigs and duration activated) of aerosol generation and exposure and determine if the system can generate rodent models with the concentrations of nicotine and cotinine resembling that in human ENDS users or other levels/PK as investigators choose for their research purposes. Our products will meet the need of the E-cig research community and greatly advance the field to enable testing potential toxicities of E- cig in animal models. Our products will be a powerful tool for studying the effects of E-cig smoking on cardiovascular, respiratory and nervous systems, on metabolism, pregnancy, development as well as teratogenicity and addiction of E-cigs. Therefore it will help inform regulators about the potential toxicity of E- cig products. We plan, in a future phase II project, to develop a commercial product line of E-cig generation and exposure systems including E-cig self-administration for rodents.